ABSTRACT New strategies to improve adoptive cell therapy (ACT) protocols are now emerging to enhance in vivo persistence of adoptively transferred tumor epitope specific T cells and overcome tumor-induced immunosuppression. Our preliminary data suggests that there is a direct correlation between the long-lived central memory T cells (Tcm) and its anti-oxidant capacity. Overexpression of thioredoxin-1 (Trx), a key molecule that regulates cell-surface thiols (c-SH), resulted in increased Tcm phenotype in T cells obtained from TCR transgenic mouse crossbred with Trx transgenic mouse, or engineering human T cells with retroviral vector with TCR and Trx together. Further, a quantification of the metabolites within Pmel vs. Pmel-Trx cells showed that metabolites from pentose-phosphate pathway (PPP) and tricarboxylic acid cycle (TCA) intermediate alpha-ketoglutarate (?-KG) were significantly higher in Pmel-Trx T cells as compared to Pmel cells. While reductive intermediates generated by PPP are important to overcome oxidative stress, recent reports have shown that ?-KG is important in extending the cellular lifespan and regulating pluripotency of stem cells. These preliminary observations lead us to hypothesize that ?the presence of Trx drives tumor reactive T cells to a c-SHhi phenotype, which not only exhibits enhanced anti-oxidant phenotype, but regulates a combination of events including post-translational modifications, and epigenetic stability that lead to metabolically fit anti-tumor T cells?. We propose the following specific aims: 1) To determine how the level of thiol/thioredoxin on the surface of T cells regulates the generation of tumor reactive Tcm/Tscm cells in vivo, 2) To determine how changes in the metabolic pathways and metabolites in T cells regulate the generation of tumor reactive Tcm/Tscm in vivo, 3) To determine if modulation of reduced thiols and/or metabolites results in generation of tumor reactive TCR transduced human T cells with functional memory phenotype. We believe that our studies are innovative and will uncover important aspects that need to be considered when generating tumor specific Tcm/Tscm cells for ACT.